The MAPKs ERK, JNK, and p38 control diverse aspects of the immune response, including regulation of cytotoxin biology in NK cells and CTL. The chemokine CCL5 is coreleased with the cytotoxins, perforin, the granzymes, and granulysin, during the lethal hit administered by cytotoxic CD8+ T cells (CTL). CCL5 expression is up-regulated relatively late in CTL coincident with their functional maturation 3-7 days after activation. Unlike T cells, NK cells have the ability to kill virally infected or transformed cells when directly isolated from the peripheral circulation. In this study, we show that in contrast to T cells, peripheral blood NK cells express CCL5 constitutively. The use of specific inhibitors of the JNK, ERK, and p38 MAPK pathways showed that the JNK pathway controls expression of CCL5 by NK cells. Promoter-reporter assays identified a compact region of the CCL5 promoter responsible for the constitutive transcription of CCL5 by NK cells. EMSA, chromatin immune precipitation, the use of heterologous promoters, and site-directed mutagenesis demonstrated that transcription in NK cells is largely controlled through binding of the transcription factor specificity protein 1 to a region -75 to -56 upstream of the site of transcriptional initiation. Specificity protein 1 expression, and in turn the constitutive expression of CCL5, was found to be controlled through constitutive activation of the JNK/MAPK pathway in peripheral blood NK cells.